Understanding the transformation of a normal cell to cancerous cell continues to be a very active area of research both for delineating the underlying molecular mechanisms involved in both genesis and maintenance of cancer as well as developing therapies which may help prevent or manage cancerous growth. While the precise molecular mechanisms leading to emergence of cancer is still being revealed, it is generally agreed that a handful of key genes which serve to regulate growth, proliferation, survival, migration and demise of cells are involved. These genes are typically of two broad classes referred to as either oncogenes or tumor suppressor genes. Both classes of genes under normal circumstances play key roles in regulating cellular processes mentioned above. However, due to certain mutations or over expression these genes are either constitutively activated (as in the case of kinases) or inactivated in the case of tumor suppressor genes such as PTEN.
While the general consensus in the field of cancer research has been that cancer is typically the result of multiple lesions that act in concert to maintain and support cancerous growth and metastasis, work over the last decade is providing evidence that at least certain kinds of cancers may depend on only a single oncogene or oncogenic pathway for growth, proliferation and survival. This hypothesis is referred to as oncogene addiction and as a corollary to this hypothesis it can be postulated targeting these key oncogenes for drug development may provide a window of opportunity for cancer treatment. Thus oncogne addiction may present the “Achilles' heel” of cancer which may be exploited therapeutically. A profound implication of this hypothesis is that switching off this crucial pathway upon which cancer cells have become dependent should have devastating effects on the cancer cell while sparing normal cells that are not similarly addicted.
Tumor dependency on the well-studied “classical” oncogenes, such as transcription factor MYC and GTPase RAS, has been demonstrated in variety of experimental models (Felsher and Bishop 1999; Wu et al. 2007; Chin et al. 1999; Fisher et al 2001).
Activated kinases have been shown to be the “Achilles' heel” of many cancers (Sharma S. V. and Settleman J. Genes Dev. 2007 21:3214-3231). A kinase is a type of enzyme that transfer phosphate groups from high-energy donor molecules, such as ATP, to specific substrates, a process referred to as phosphorylation. One of the largest groups of kinases are protein kinases, which act on and modify the activity of specific proteins. More than 500 different protein kinases have been identified in human; of this 11% are known to be receptor tyrosine kinases (RTKs). Various other kinase act on small molecules such as lipids, carbohydrates, amino acids and nucleotides, either for signaling or prime them for metabolic pathway. In addition to the functions in normal tissues/organs, many kinses also play more specialized roles in a host of human diseases including cancer. A subset of kinases (also referred to as oncogenic kinases), when dysregulated, can cause tumor formation/growth and further contribute to tumor maintenance and progression. Thus, oncogenic kinases represent one of the largest and most attractive groups of targets for cancer intervention and drug development.
ABL and platelet-derived growth factor receptor (PDGFR) tyrosine kinase, which are targets of imatinib, are often activated by chromosomal translocations (BCR-ABL, TEL-ABL, TEL-PDGFR). Tumor cell lines harboring these activated ABL and PDGFR become addicted to them for their survival and undergo apoptosis following inactivation of these two concogenes. The clinical success of imatinib in treating chronic myelogenous leukemia (CML) and gastrointestinal stromal tumor (GIST) is the first examples of oncogene addiction in the context of cancer therapy. Imatinib, which also inhibits the KIT receptor tyrosine kinase, cause apoptosis of small cell lung cancer (SCLC) cell lines addicted to the autocrine loop created by the expression of KIT as well as its ligand, stem cell factor in these cells in culture or xenografts. Additionally, mutations in KIT in GIST renders these cells addicted to the KIT oncoprotein, and its inactivation leads to apoptosis of the tumor cells.
Oncogene addiction also contributes to the clinal success of agents that target HER2. The HER2 oncogene is amplified in 25-30% of breast cancers, suggesting that these tumors may be addicated to HER2. Consistent with this hypothesis, breast cancer cells in culture or grown as xenografts are preferentially growth inhibited by HER2 inhibition. These finding led to the clinical success of HER2 targeted antibodies, Trastuzumab/Herceptin and Pertuzumab in treatment of patients with HER2-positive metastatic breast cancer.
The use of selective epidermal growth factor receptor (EGFR) kinase inhibitors in lung cancer treatment presents another example of onocogene addiction that has yielded clinical success. Mutations of the kinase domain of EGFR are found in a 10-20% non-small cell lung cancer (NSCLC), and significant clinical responses to EGFR inhibitors (gefitinib and erlotinib) have been well correlated to such mutations. Glioblastomas harboring EGFR gene amplification and deletion mutations appear to be addicted to these EGFR activating mutations.
The use of mutant specific b-Raf (V600E) inhibitor (Vemurafenib/PLX4032) in treatment of late-stage melanoma presents another example of onocogene addiction that has yielded clinical success. About 60% of melanomas have V600E mutation. PLX4032 has been shown to cause apoptosis in these melanoma cell lines (Hatzivassiliou, et al. Nature 2010 464:431-5). And the growth of a melanoma cell line A375 has been shown to be inhibited by silencing the bRAF gene by short hairpin RNA (Sala, et al. Mol. Cancer Res. 2008 6:751-9).
The use of ALK kinase inhibitor in NSLC treatment is another clinical success utilizing oncogene addiction concept. About 4% of patients with NSCLC have a chromosomal rearrangement that generates a fusion gene between EML4 (echinoderm microtubule-associated protein-like 4) and ALK (anaplastic Lymphoma kinase) and about 60% of Anaplastic Large Cell Lymphomas (ALCL) have a chromosomal translocation that results a fusion gene between NPM (nucloplasmin) and ALK. Both fusions result in constitutive kinase activity that contributes to carcinogenesis and seems to drive the malignant phenotype. ALK mutations are also thought to be important in driving the malignant phenotype in about 15% of cases of neuroblastoma, a rare form of central neverous system cancer that occurs almost exclusively in very young children. Crizotinib/PF02341066 has successfully shown to cause tumor shrinkage or stabilizing disease in 90% of patients carrying the ALK fusion gene (Hem Onc Today 2010-06-05).
In addition to the clinical successes of a few kinase inhibitors to which tumor cells have become addicted, more clinical data indicates that this phenomenon may be apply to a large number of other kinases. For examples, MET gene amplifications, as well mutations and abnormal expression of the MET signaling pathway have been observed in a significant fraction of gastric cancers, lung cancers and prostate cancers. The fibroblast growth factor receptor 3 (FGFR3) is activated in 15% of multiple myelomas by chromosomal translocation. Aurora kinases are frequently amplified in a diverse array of human cancers such as leukemia, colon and pancreatic tumors. Genetic aberrations of PI3K, which lead to constitutive activation, are commonly observed in human cancers (Bader et al. Nature Review 2005 5: 921-9). Lastly, the RET onocogene is frequently mutated in medullary thyroid carcinomas and subset of papillary thyroid cancers. It has been shown that inactivation of these mutated kinase by a variety of methods in different systems typically results in growth inhibition of tumor cell death.
Recent studies have shown that additional classes of genes that may also confer a state of dependency in cancer when dysregulated. For example, oncogenic RNAs (“oncomirs”) have emerged as important players in cancer. The role of oncomirs in oncogene addition is demonstrated by the fact that antisense inhibition of these oncomirs led to apoptosis of lunger cancer cells overexpressing the corresponding oncomirs (Matsubara et al. Oncogene 2007 26: 6099-6105.)
In order to develop therapies for targeting key oncogenes involved in cancer, it is important to establish both in vitro and in vivo models that can be used for screening and evaluation of lead compounds. One of the advantages for developing in vitro oncogene addiction models is that certain cancer cell lines continue to maintain the oncogene addiction state even when cultured in petri-dishes or microtiter plates. The oncogene addiction status of these cell lines can be evaluated by using tool compounds or other reagents which inhibit the oncogene and typically results in cytostasis or apoptosis. Various molecular biological or cellular biological methods could be used to assay or evaluate the response of these oncogene addicted cells to various tool compounds or other reagents.